Pancreatic β cells secrete insulin in response to elevated amounts of blood glucose and other blood borne metabolites termed “secretagogues.” Glucose is delivered to body tissues in part via transporters that respond to insulin. Insulin reduces blood glucose by stimulating its removal from the bloodstream and by inhibiting glucose production by the liver. In addition to glucose, other molecules stimulate β-cells to secrete insulin. These include amino acids, α-ketoacids and mitochondrial metabolites. Collectively, these secondary secretagogues can account for as much as 50% of all insulin secreted.
The control of blood glucose is initiated by Glut2-dependent glucose uptake by pancreatic β-cells. Inside the cell, glucose is metabolized to yield ATP, which causes ATP-sensitive K+ion channels (KATP) to close, blocking the efflux of K+ ions and depolarizing the membrane potential. This depolarization opens voltage-gated Ca2+ ion channels (VCa), leading to a rise in intracellular Ca2+, which triggers vesicle mobilization and insulin secretion.
A genetically inherited condition characterized by higher than normal glutamate dehydrogenase enzyme activity arises from any of several mutations at the site of GTP-mediated suppression of the enzyme. The enzyme catalyzes oxidative glutamate deamination and produces ammonia, α-ketoglutarate (αKG), and reducing equivalents. Humans having the condition exhibit hyperammonemia, hyperinsulinemia, and hypoglycemia. The mechanism by which excessive glutamate dehydrogenase activity increases insulin secretion is not known. The conventional wisdom is that stimulation of insulin release requires metabolism of αKG.
αKG is a Krebs cycle intermediate that can be oxidatively decarboxylated to form succinate, an insulin secretagogue. αKG is also a stoichiometric cofactor of various αKG-dependent hydroxylase enzymes, including prolyl-4-hydroxylases which have a variety of substrates, the best studied being collagen. Prolyl-4-hydroxylases also regulate hydroxylation at a single proline residue on Hypoxia Inducible Factor-1α(HIF-1α), a transcription factor that regulates a potassium ATP (KATP) channel involved in insulin secretion from β cells.
Cellular αKG is formed by glutamate dehydrogenase (GDH) and in the citric acid cycle via isocitrate dehydrogenase as well as by the branched chain aminotransferase (BCAT) reaction wherein no reducing equivalents are produced. In this reaction, an α-ketoacid and an α-amino acid are interconverted into their corresponding amino- and α-ketoacids. α-ketoisocaproic acid (KIC) is transaminated using an amine group from glutamate to form leucine. In the process, glutamate loses an amine group and forms αKG. KIC-induced hypersecretion of insulin is prevented by blocking the transamination of KIC (using, e.g., BCAT inhibitor methyl-leucine) to leucine and the attendant formation of αKG. αKG formation may be part of a signaling cascade that leads to chronic insulin hypersecretion. When applied to isolated pancreatic β-cells, KIC is known to cause depolarization of the cell membrane voltage, leading to generation of action potentials, increase in cytosolic Ca2+ ion concentration and increased insulin secretion. KIC-dependent depolarization is known to be due to a direct inhibition of KATP. In intact β-cells the flow of K+ ions through KATP is likely the current that dominates the resting membrane potential. Accordingly, agents that modulate KATP channel activity will necessarily alter membrane potential.
Persaud, S. J. et al., J. Molec. Endocrin. 22:19-28 (1999) discloses that protein tyrosine kinase inhibitors genistein and tyrphostin A47 inhibited KIC-stimulated insulin release without affecting glucose metabolism. Persaud et al. suggested that the protein tyrosine kinase inhibitors exert their inhibitory effects distal to closure of ATP-sensitive K+ channels, but proximal to Ca2+ entry into β cells, and that the inhibitors act at the site of the voltage-dependent Ca2+ channel that regulates Ca2+ influx into β cells following depolarization.
A recently discovered heritable genetic disorder attributable to mutations in the GDH enzyme highlights the physiological importance of αKG-dependent insulin segretagogue activity. The enzyme catalyzes conversion of glutamate to αKG and ammonia. In mutated form, the enzyme causes chronic hyper-insulinemia resulting in severe hypoglycemia. Interestingly, the mutations led to an increase in enzyme activity and over-production of αKG.
It would be advantageous to identify compounds that augment the amount of insulin secretion evoked by glucose and other secretagogues. Such compounds would have therapeutic utility in treating those forms of diabetes caused by insufficient β cell responsiveness to insulin secretagogues.